PROJECT SUMMARY/ABSTRACT Hexavalent chromium [Cr(VI)] exposure causes multiple toxic effects in humans. The main concern of Cr(VI) toxicity is its carcinogenicity as Cr(VI) is one of the most well-recognized environmental and occupational carcinogens causing lung and other cancer in humans. While it is under active study, the mechanism of Cr(VI) carcinogenicity remains elusive. Previous studies showed that Cr(VI) exposure causes genotoxic effects, which are thought to play important roles in Cr(VI) carcinogenicity. On the other hand, studies also showed that Cr(VI) exposure dysregulates epigenetics such as increased DNA methylation and abnormal histone posttranslational modifications. However, it remains to be determined how dysregulated epigenetics contributes to Cr(VI) carcinogenicity and if Cr(VI)-caused epigenetic changes play a role in its genotoxic effect. Our preliminary studies showed: (i) Chronic Cr(VI) exposure increases the levels of histone H3 repressive methylation marks (H3K9me2 and H3K27me3) and their related histone methyltransferases (HMTases) (G9a, SUV39H1, and EZH2). (ii) Up- regulation of HMTases play a causal role in Cr(VI)-induced cancer stem cell (CSC)-like property and cell malignant transformation. (iii) Chronic Cr(VI) exposure down-regulates the expression of O6-methylguanine DNA methyltransferase (MGMT), a key gene in DNA repair network; and MGMT down-regulation plays a causal role in Cr(VI)-induced cell transformation. (iv) The level of O6-methylguanine (O6-MeG), a highly mutagenic DNA lesion, is significantly increased in Cr(VI)-transformed cells; but stably expressing MGMT greatly reduces Cr(VI) exposure-induced O6-MeG. (v) Cr(VI) exposure also significantly decreases the level of MGMT but increases O6-MeG level in mouse and human lung tissues. (vi) Stably knocking down HMTases G9a or SUV39H1 significantly increases MGMT level but reduces O6-MeG level in Cr(VI)-exposed cells. (vii) Cr(VI)- transformed cells display impaired DNA damage repair capacity but stably expressing MGMT significantly reduces chronic Cr(VI) exposure-caused DNA damage repair deficiency. And (viii) Treatment with a natural compound dihydromethysticin (DHM) is capable of increasing MGMT level and reducing O6-MeG level in Cr(VI)- transformed cells. (viii) We generated a conditional and lung specific MGMT expression transgenic mouse model. Based on these findings from our preliminary studies and that from literature reports, our central hypothesis is that up-regulation of HMTases by chronic Cr(VI) exposure down-regulates the expression of MGMT leading to increased level of highly mutagenic DNA lesion O6-MeG and promoting Cr(VI) carcinogenesis. Three aims are proposed: Aim 1 will determine the mechanism by which chronic Cr(VI) exposure causes MGMT down- regulation. Aim 2 will determine the role of MGMT in C(VI)-induced lung tumorigenesis using a conditional and lung specific MGMT expression transgenic mouse model. And Aim 3 will determine the protective effect of the natural compound DHM treatment on Cr(VI)-induced cell transformation and tumorigenesis in mice.